We will develop a novel system for the study of cell fusion, testing two different fusion- activated reporter-gene systems to quantify newly fused myogenic and osteoclastogenic cells in culture and in the developing mouse. Both the lacZ complementation and Cre-lox recombination systems provide the opportunity to readily score cell fusion activity in any cell type in vivo or in vitro. In each system, a pair of distinct mouse strains will be used: - For the lacZ complementation system, we will create new transgenic strains, each expressing one of two inactive proteins. Upon mixing of fused cell cytoplasms, these mutant proteins reconstitute active beta-galactoside activity. - For the Cre-lox-GFP system, we will use one mouse harboring a silent, Cre-activatable lacZ or GFP gene, and another with ubiquitously expressed Cre recombinase. Upon mixing of fused cell contents, Cre recombinase will rearrange and activate expression of the reporter gene. In either system, when cells from two complementary transgenic strains are combined,-in vitro pr within chimeric mice - cell fusion events will result in activation of the reporter. Only syncytia formed by fusion will express the reporter activity, and they will thus be readily detectable either by microscopy, FACS analysis, or quantitative assay of lysates. Using such a simple scheme, developed and validated during this pilot project, we will in the future be able to perform a wide range of experiments to understand the developmental regulation, and cell-biological and molecular mechanisms of cell membrane fusion in myogenesis and osteoclastogenesis. To explore the potential of this system, we will purchase the assistance of the UCHC transgenic mouse facility in creation of transgenic mice and aggregation and implantation of chimeric mouse embryos; we will then apply standard techniques of myoblast and osteoclast cell culture to validate the system in vitro. Our pilot-phase analysis of these systems will rely heavily on the Imaging Core of the CCMD.